Anisomeles (Family: Lamiaceae) is a genus of herbaceous or shrubby plants distributed from Africa to India, South East Asia to North East Australia and east from China to Philippines. Genus Anisomeles is represented in Maharashtra by three species viz. A. heyneana, A. malabarica and A. indica of which A. heyneana is endemic to Maharashtra. A. malabarica and A. indica have been examined chemically and found to contain cembrane diterpenes, triterpenes, flavonoids, etc. while there is no reported phytochemical work on A. heyneana. 
High Performance Thin Layer Chromatography (HPTLC) method has been developed for the quantification of quercetin, β-sitosterol, stigmasterol, catechin and ovatodiolide present in A. indica and A. malabarica. 
Anisomeles indica is used in folk medicine all over the India, where it has received widespread use as folk medicine, predominantly in the treatment of intestinal disorders and intermittent fever. Anisomeles indica has anti-microbial, astringent, carminative properties. Ethanolic extract (50%) of the herb showed hypothermic activity and when burnt, acted as a mosquito repellant. The essential oil present in the herb is useful in uterine affections, whereas, Anisomeles malabarica is useful in halitosis, epilepsy, hysteria, amnesia, anorexia, dyspepsia, colic, flatulence, intestinal worms, fever arising from teething children, intermittent fever, gout, swelling and diarrhea.
Anisomeles heyneana, commonly known as western hill catmint, is a tall, erect herb with slender stems with branches quadrangular having oppositely arranged ovate lance-like leaves. The flowers are white, tinged with pink, and 2-lipped.
Tuberculosis is a major and still neglected cause of death and disability with around 2 million deaths worldwide in 2009 and 9 million infections. There is more tuberculosis today than at any other time in History. The emergence of drug resistant strains and confluence with HIV epidemic has turned TB into a global public health crisis. Although, available drug regimens can cure most patients, emergence of MDR, SDR and XDR-TB coupled with insufficient global drug pipeline, justifies continued efforts towards development of new drugs with new mode of action and novel structures.
Taiwan Publication No. 201023966 employs water decoction cooking method and modern scientific isolation and purification technologies, with Anisomeles indica (L.) Kuntze stem as extraction material, to derive effective ingredients. However, Taiwan Application No. 20100120578 provides composition that contains ovatodiolide with effective dosage to inhibit Helicobacter pylori growth, wherein ovatodiolide is derived from dried leaves of Anisomeles indica (L.) Kuntze.
The extraction/isolation of cembrane type diterpenoid compounds from Anisomeles indica are known in the prior art; such as two cembrane type diterpenoid compounds, namely ovatodiolide and 4,5 epoxovatodiolide isolated from Anisomeles indica, which exhibits selective antiplatelet aggregation activities toward collagen is disclosed in Journal of Natural Products 2008 July 71(7), 1207-1212, whereas the isolation of cembrane-type diterpenoid (ovatodiolide) extracted from the whole plants of Anisomeles indica and their anti-inflammatory activities is reported by Yerra Rao et al. in Journal of Ethnopharmacology 121, (2), 292-296. Biologically active macrocyclic diterpenoids such as 4,7-oxycycloanisomellic acid, 4-methylene-5-hydroxyovatodiolode and 4-methylene-5-oxoanisomelic acid from (Anisomeles indica,) together with ovatodiolide, 4-5 epoxyovatodiolide and anisomelic acid having cytotoxic activity and calcium antagonistic activity is described in Plant Med 1986; 52(1); 38-41. However extraction of ovatodiolide and other diterpenoids from dried aerial parts of Anisomeles ovata is reported in Journal of Natural Products 47, (6), 1052-1068, 1984. Work by Shahidul Alam et al. in Elsevier Fitoterapia, September 2000 71(5):574-6 relates to ovatodiolide, from Anisomeles indica and its anti-HIV activity.
Formulation containing macrocyclic diterpene-ovatodiolide isolated from the roots of Anisomeles malabarica is known in the art. Further the isolation of diterpenoid [16,17-dihydroxy-16beta(−)kauran-19-oic acid] belongs to kaurane derivative from Beyeria species (Family: Euphorbiaceae) is disclosed by Fujita, Eiichi et al in Bulletin of the Institute for Chemical Research, Kyoto University (1966), 44(3): 239-272; whereas a oxygenated rare phyllocladane like phyllocladan-16a,19-diol along with flavonoids such as kaempferol-3-O-β-D-galactopyranoside, kaempferol-3-O-L-rhamnopyranoside and scopoletin, from the leaves of Ailanthus triphysa (Family: Simaroubaceae) is reported in Chinese Journal of Chemistry” 21, 2200-203, February 2003. Further the 13β-kaurane from Plectranthus ambiguus as inhibitors of the arachidonate metabolism and allergens is disclosed in Helvetica Chimica Acta-Vol. 86 (2003).
Leukemia is often referred to as cancer of the blood. It is characterized by the widespread uncontrolled proliferation of large numbers of abnormal blood cells, usually white blood cells, which take over the bone marrow and quickly spread to the blood stream. Other organs that may also be affected include lymph nodes, spleen, liver, central nervous system and other organs. As of 2010 there were an estimated 43,050 new cases of leukemia and 21,840 deaths due to leukemia. Everyday 118 people are diagnosed with leukemia and 60 of those lose the fight (http://www.leukemia-research.org/page.aspx?pid=214.
THP1 (Human acute monocytic leukemia cell line) is a well known model cell line to assess cytotoxic effects of drug candidates.
International Journal of Cancer Research 2007, 3 (4), 174-179 discloses study of the anticancer effect of crude ethanolic leaf extract of Anisomeles malabarica, which is evaluated on Diethylnitrosamine (DEN) in mice. The assessment of anticancer activity is evaluated by measuring the activities of total protein, Glutamate Pyruvate Transaminase (GPT), Glutamate Oxaloacetate Transaminase (GOT), Acid Phosphatase (ACP) and Alkaline Phosphatase (ALP) wherein the ethanolic extract at an oral dose of 100 mg kg−1 is found to exhibit a significant (p<0.05) protective effect by reducing liver and serum levels of total protein, GPT, GOT, ACP and ALP as compared to DEN induced mice.
With regards to photosensitization technique, the solar energy is the most abundant, clean and green source of renewable energy and it can be harnessed via different routes in the interest of diverse applications. In many applications such as dye-sensitized solar cells (DSSCs), photocatalysis, solar to chemical conversion, etc., sensitizers with good optical properties over the visible range are essential. In the case of DSSCs, a number of factors such as the choice of the photoanode, the dye, the bottom, the counter electrodes, etc., can limit the cell performance, and a quantitatively significant stable electronic anchorage of the dye to the surface of the metal oxide nanostructure (e.g., TiO2, ZnO) is a critical factor that determines the cell efficiency. Among the various dyes that have been tested as sensitizers, ruthenium-based dyes have given the highest efficiency of ˜11% on the laboratory scale. The main advantage of Ru-based dyes is their efficient metal to ligand charge transfer.
However, their high cost due to complex synthetic procedures as well as environmental hazards posed by the presence of heavy metals invite search for safer and cheaper alternatives. There have been interesting explorations of natural dyes in the context of the dye-sensitized solar cell (DSSC) application using pigments obtained from biomaterials such as flowers, fruits and leaves. The natural pigments commonly extracted from flowers and fruits are anthocyanins. Anthocyanins give colors ranging from red to blue depending on the pH of the medium. The maximum efficiency obtained by use of anthocyanins, i.e., juice obtained from Hibiscus sabdariffa, is 3.1%. The problem with anthocyanins is that they are pH sensitive, i.e., good binding with TiO2 occurs only if anthocyanin is present as flavylium ion species (which is stable) at pH around 1 to 3.2, but if pH is increased this ion gets hydrated to form quinoidal bases. These quinoidal compounds are labile and can be transformed into colorless compounds. If pH becomes more acidic <1 then the compound itself is leached out. Maintaining the right pH is nontrivial and at the same time a very crucial step while extracting anthocyanins. These anthocyanins are also thermally unstable. Porphyrins, which form the structural core of chlorophylls, the natural light harvesters, have also been examined as another class of sensitizers for DSSCs. Campbell et al. have synthesized metal-porphyrin dyes that show conversion efficiency of 7.1%. In 1994, M. Gratzel studied the mechanism of photosensitization of TiO2 solar cells by chlorophyll derivatives whereas, Kamat and co-workers in 1986 also explored the use of a chlorophyll analogue as a sensitizer for DSSCs. Natural chlorophyll extracted from Shiso leaves was used as sensitizer by Kumara et al., which showed efficiency of 0.59%. Isolated chlorophyll was used along with shisosin (an anthocyanin) to realize a synergistic effect that gave efficiency of 1.31%. Betalain pigments extracted from red beet root were also used as dyes sensitizer which gave efficiency of 0.19%. More recently, Wang and co-workers have done interesting studies on the chlorophyll system and have also modified the dye with metal conjugation to enhance efficiency.
Recenly, work by Shruti et al. in ACS Applied Materials and Interfaces (dx.doi.org/10.1021/am200341y) has demonstrated Isobutrin as a promising dye sensitizer in DSSCs. It belongs to chalcone class of natural products and in this work this class was reported first time as having potential to be developed as dye sensitizer.
Although a number of molecular sensitizers in the form of organic dyes (including heavy metal ion incorporated ones) have been synthesized and are being used, the corresponding methods of synthesis, application and prolonged use are not eco-friendly, cost-effective, with photo-, thermal-degradability dramatically reducing half life of sensitizer and hence affecting the coast and they may not be stable. It is therefore highly desirable to explore sources of new natural dye systems that are stable, nontoxic (biocompatible), and with the desirable optoelectronic properties.
In view of above drivers, uses and necessities of exploring natural products to supply respective leads, natural products like ovatodiolide, phyllocladane as well as phenolic compounds like verbascoside having different biological and physical properties can be isolated from different herbaceous plants. The existing diterpene compounds isolated from herbaceous plants do not exhibit such diverse applications as in the field of pharmaceutical, dye and solar energy.
Hence there is need to isolate and evaluate diterpenoids from the extract of Anisomeles species having biological activity such as anti-mycobacterial, anti-proliferative activity and solar cell sensitizing activity.
With due experiments the present inventors have succeeded to evaluate diterpenoids belonging to cembrane and phyllocladane class isolated from aerial parts of Anisomeles heyneana for significant anti-mycobacterial activity against M. tuberculosis and anti-proliferative activity against human Thp-1 cell line respectively.
The invention further provides another phenethylglycoside compound, verbascoside isolated from same species found to be effective dye-sensitizer in dye sensitized-solar cells (DSSCs).
Therefore, the objective of the present invention is to provide process for the isolation of non-toxic, economically viable and stable diterpenoids and phenethylglycoside isolated from extract of Anisomeles heyneana. 